Evolution of the karyopherin-β family of nucleocytoplasmic transport factors; ancient origins and continued specialization

Background

Macromolecular transport across the nuclear envelope (NE) is achieved through nuclear pore complexes (NPCs) and requires karyopherin-βs (KAP-βs), a family of soluble receptors, for recognition of embedded transport signals within cargo. We recently demonstrated, through proteomic analysis of trypanosomes, that NPC architecture is likely highly conserved across the Eukaryota, which in turn suggests conservation of the transport mechanisms. To determine if KAP-β diversity was similarly established early in eukaryotic evolution or if it was subsequently layered onto a conserved NPC, we chose to identify KAP-β sequences in a diverse range of eukaryotes and to investigate their evolutionary history.

Results

Thirty six predicted proteomes were scanned for candidate KAP-β family members. These resulting sequences were resolved into fifteen KAP-β subfamilies which, due to broad supergroup representation, were most likely represented in the last eukaryotic common ancestor (LECA). Candidate members of each KAP-β subfamily were found in all eukaryotic supergroups, except XPO6, which is absent from Archaeplastida. Phylogenetic reconstruction revealed the likely evolutionary relationships between these different subfamilies. Many species contain more than one representative of each KAP-β subfamily; many duplications are apparently taxon-specific but others result from duplications occurring earlier in eukaryotic history.

Conclusions

At least fifteen KAP-β subfamilies were established early in eukaryote evolution and likely before the LECA. In addition we identified expansions at multiple stages within eukaryote evolution, including a multicellular plant-specific KAP-β, together with frequent secondary losses. Taken with evidence for early establishment of NPC architecture, these data demonstrate that multiple pathways for nucleocytoplasmic transport were established prior to the radiation of modern eukaryotes but that selective pressure continues to sculpt the KAP-β family.     

Depletion of nucleoporins from HeLa nuclear pore complexes to facilitate the production of ghost pores for in vitro reconstitution

During cell division, Nuclear Pore Complexes (NPCs) are broken down into protein subcomplexes that are the basis for reassembly in daughter cells. This is the driving force for the establishment of an in vitro reconstitution system to study aspects of NPC reassembly. In this study, nuclear envelope (NE) was isolated from HeLa cells. NE was treated with increasing concentrations of heparin to extract nucleoporins (Nups) for the production of “ghost pores” which are pores severely deficient in Nups, while still containing Pore Membrane proteins (POM) needed to anchor the NPC. Ghost pores have been subjected to incubation with previously stripped Nups and some re-binding has been shown to occur by western blot analysis. This in vitro assay provides a powerful tool to investigate the protein–protein interactions of NPC reassembly from a human cell line. Through a better understanding of the process of NPC reassembly, we can continue to piece together the puzzle of this macromolecular structure. It is most advantageous to establish a straightforward reconstitution procedure at the mammalian level.     

Evaluation of Human Leukocyte Antigen-A (HLA-A), Other Non-HLA Markers on Chromosome 6p21 and Risk of Nasopharyngeal Carcinoma

Background

The association between human leukocyte antigen (HLA) genes (located in the Major Histocompatibility Complex [MHC] region of chromosome 6p21) and NPC has been known for some time. Recently, two genome-wide association studies (GWAS) conducted in Taiwan and China confirmed that the strongest evidence for NPC association was mapped to the MHC region. It is still unclear, however, whether these findings reflect direct associations with Human Leukocyte Antigen (HLA) genes and/or to other genes in this gene-rich region.

Methods

To better understand genetic associations for NPC within the MHC region of chromosome 6, we conducted an evaluation that pooled two previously conducted NPC case-control studies in Taiwan (N = 591 cases and N = 521 controls). PCR-based genotyping was performed for 12 significant SNPs identified within 6p21 in the Taiwan NPC GWAS and for the HLA-A gene (exons 2 and 3).

Findings

After confirming homogeneity between the two studies, pooled odds ratios (OR) and 95% confidence intervals (CI) were estimated by logistic regression. We found that HLA-A (p-trend = 0.0006) and rs29232 (within the GABBR1 gene; p-trend = 0.005) were independent risk factors for NPC after adjustment for age, gender, study and each other. NPC risk was highest among individuals who were homozygous for the HLA-A*0207 risk allele and carriers of the rs29232 risk allele (A).

Conclusion

Our study suggests that most of the SNPs significantly associated with NPC from GWAS reflect previously identified HLA-A associations. An independent effect of rs29232 (GABBR1), however, remained, suggesting that additional genes within this region might be associated with NPC risk.     

Pom33, a novel transmembrane nucleoporin required for proper nuclear pore complex distribution

The biogenesis of nuclear pore complexes (NPCs) represents a paradigm for the assembly of high-complexity macromolecular structures. So far, only three integral pore membrane proteins are known to function redundantly in NPC anchoring within the nuclear envelope. Here, we describe the identification and functional characterization of Pom33, a novel transmembrane protein dynamically associated with budding yeast NPCs. Pom33 becomes critical for yeast viability in the absence of a functional Nup84 complex or Ndc1 interaction network, which are two core NPC subcomplexes, and associates with the reticulon Rtn1. Moreover, POM33 loss of function impairs NPC distribution, a readout for a subset of genes required for pore biogenesis, including members of the Nup84 complex and RTN1. Consistently, we show that Pom33 is required for normal NPC density in the daughter nucleus and for proper NPC biogenesis and/or stability in the absence of Nup170. We hypothesize that, by modifying or stabilizing the nuclear envelope–NPC interface, Pom33 may contribute to proper distribution and/or efficient assembly of nuclear pores.     

The structure of the NPC1L1 N-terminal domain in a closed conformation

Background

NPC1L1 is the molecular target of the cholesterol lowering drug Ezetimibe and mediates the intestinal absorption of cholesterol. Inhibition or deletion of NPC1L1 reduces intestinal cholesterol absorption, resulting in reduction of plasma cholesterol levels.

Principal Findings

Here we present the 2.8 Å crystal structure of the N-terminal domain (NTD) of NPC1L1 in the absence of cholesterol. The structure, combined with biochemical data, reveals the mechanism of cholesterol selectivity of NPC1L1. Comparison to the cholesterol free and bound structures of NPC1(NTD) reveals that NPC1L1(NTD) is in a closed conformation and the sterol binding pocket is occluded from solvent.

Conclusion

The structure of NPC1L1(NTD) reveals a degree of flexibility surrounding the entrance to the sterol binding pocket, suggesting a gating mechanism that relies on multiple movements around the entrance to the sterol binding pocket.     

Non-Gaussian Analysis of Diffusion Weighted Imaging in Head and Neck at 3T: A Pilot Study in Patients with Nasopharyngeal Carcinoma

Purpose

To technically investigate the non-Gaussian diffusion of head and neck diffusion weighted imaging (DWI) at 3 Tesla and compare advanced non-Gaussian diffusion models, including diffusion kurtosis imaging (DKI), stretched-exponential model (SEM), intravoxel incoherent motion (IVIM) and statistical model in the patients with nasopharyngeal carcinoma (NPC).

Materials and Methods

After ethics approval was granted, 16 patients with NPC were examined using DWI performed at 3T employing an extended b-value range from 0 to 1500 s/mm². DWI signals were fitted to the mono-exponential and non-Gaussian diffusion models on primary tumor, metastatic node, spinal cord and muscle. Non-Gaussian parameter maps were generated and compared to apparent diffusion coefficient (ADC) maps in NPC.

Results

Diffusion in NPC exhibited non-Gaussian behavior at the extended b-value range. Non-Gaussian models achieved significantly better fitting of DWI signal than the mono-exponential model. Non-Gaussian diffusion coefficients were substantially different from mono-exponential ADC both in magnitude and histogram distribution.

Conclusion

Non-Gaussian diffusivity in head and neck tissues and NPC lesions could be assessed by using non-Gaussian diffusion models. Non-Gaussian DWI analysis may reveal additional tissue properties beyond ADC and holds potentials to be used as a complementary tool for NPC characterization.     

Insight into structure and assembly of the nuclear pore complex by utilizing the genome of a eukaryotic thermophile

Despite decades of research, the structure and assembly of the nuclear pore complex (NPC), which is composed of ～30 nucleoporins (Nups), remain elusive. Here, we report the genome of the thermophilic fungus Chaetomium thermophilum (ct) and identify the complete repertoire of Nups therein. The thermophilic proteins show improved properties for structural and biochemical studies compared to their mesophilic counterparts, and purified ctNups enabled the reconstitution of the inner pore ring module that spans the width of the NPC from the anchoring membrane to the central transport channel. This module is composed of two large Nups, Nup192 and Nup170, which are flexibly bridged by short linear motifs made up of linker Nups, Nic96 and Nup53. This assembly illustrates how Nup interactions can generate structural plasticity within the NPC scaffold. Our findings therefore demonstrate the utility of the genome of a thermophilic eukaryote for studying complex molecular machines.     

Reduced CTGF Expression Promotes Cell Growth,Migration, and Invasion in Nasopharyngeal Carcinoma

Background

The role of CTGF varies in different types of cancer. The purpose of this study is to investigate the involvement of CTGF in tumor progression and prognosis of human nasopharyngeal carcinoma (NPC).

Experimental design

CTGF expression levels were examined in NPC tissues and cells, nasopharynx (NP) tissues, and NP69 cells. The effects and molecular mechanisms of CTGF expression on cell proliferation, migration, invasion, and cell cycle were also explored.

Results

NPC cells exhibited decreased mRNA expression of CTGF compared to immortalized human nasopharyngeal epithelial cell line NP69. Similarly, CTGF was observed to be downregulated in NPC compared to normal tissues at mRNA and protein levels. Furthermore, reduced CTGF was negatively associated with the progression of NPC. Knocking down CTGF expression enhanced the colony formation, cell migration, invasion, and G1/S cell cycle transition. Mechanistic analysis revealed that CTGF suppression activated FAK/PI3K/AKT and its downstream signals regulating the cell cycle, epithelial-mesenchymal transition (EMT) and MMPs. Finally, DNA methylation microarray revealed a lack of hypermethylation at the CTGF promoter, suggesting other mechanisms are associated with suppression of CTGF in NPC.

Conclusion

Our study demonstrates that reduced expression of CTGF promoted cell proliferation, migration, invasion and cell cycle progression through FAK/PI3K/AKT, EMT and MMP pathways in NPC.     

Potential tumor suppressor NESG1 as an unfavorable prognosis factor in nasopharyngeal carcinoma

Background

Recently we identified nasopharyngeal epithelium specific protein 1 (NESG1) as a potential tumor suppressor in nasopharyngeal carcinoma (NPC). The purpose of this study is to investigate the involvement of NESG1 in tumor progression and prognosis of human NPC.

Methodology/Principal Findings

NESG1 protein expression in NPC was examined. Survival analysis was performed using Kaplan-Meier method. The effect of NESG1 on cell proliferation, migration, and invasion were also investigated.

Results

NESG1 expression was downregulated in atypical hyperplasia and NPC samples compared to normal and squamous nasopharynx tissues. Reduced protein expression was negatively associated with the status of NPC progression. Patients with lower NESG1 expression had a shorter overall survival and disease-free time than did patients with higher NESG1 expression. Multivariate analysis suggested NESG1 expression as an independent prognostic indicator for NPC patient survival. Proliferation, migration, and invasion ability were significantly increased in cell lines following lentiviral-mediated shRNA suppression of NESG1 expression. Microarray analysis indicated that NESG1 participated in multiple pathways, including MAPK signaling and cell cycle regulation. Finally, DNA methylation microarray examination revealed a lack of hypermethylation at the NESG1 promoter, suggesting other mechanisms are involved in suppressing NESG1 expression in NPC.

Conclusion

Our studies are the first to demonstrate that decreased NESG1 expression is an unfavorable prognostic factor for NPC.     

Can Global Variation of Nasopharynx Cancer Be Retrieved from the Combined Analyses of IARC Cancer Information (CIN) Databases?

Background

The international nasopharynx cancer (NPC) burdens are masked due to the lack of integrated studies that examine epidemiological data based on up-to-date international disease databases such as the Cancer Information (CIN) databases provided by the International Agency for Research on Cancer (IARC).

Methods

By analyzing the most recently updated NPC epidemiological data available from IARC, we tried to retrieve the worldwide NPC burden and patterns from combined analysis with GLOBOCAN2008 and the Cancer Incidence in Five Continents (CI5) databases. We provide age-standardized rates (ASR) for NPC mortality in 20 highest cancer registries from GLOBOCAN2008 and the World Health Organization (WHO) mortality databases, respectively. However, NPC incidence data can not be retrieved since it is not individually listed in CI5 database. The trend of NPC mortality was investigated with Joinpoint analysis in the selected countries/regions with high ASR.

Results

GLOBOCAN 2008 revealed that the highest NPC incidence rates in 2008 were in registries from South-Eastern Asia, Micronesia and Southern Africa with Malaysia, Indonesia and Singapore ranking the top 3. WHO mortality database analysis revealed that China Hong Kong, Singapore and Malta ranks the top 3 regions with the highest 5-year mortality rates.

Conclusions

NPC mortality rate is about 2–3 times higher in male than that in female, and shows decrease tendency in those selected countries/regions during the analyzed periods. However, the integrated analyses of the current IARC CIN databases may not be suitable to retrieve epidemiological data of NPC. Much effort is required to improve the local cancer entry and regional death-reporting systems so as to aid similar studies.     

The nucleoporin Nup188 controls passage of membrane proteins across the nuclear pore complex

All transport across the nuclear envelope (NE) is mediated by nuclear pore complexes (NPCs). Despite their enormous size, ∼60 MD in vertebrates, they are comprised of only ∼30 distinct proteins (nucleoporins or Nups), many of which form subcomplexes that act as building blocks for NPC assembly. One of these evolutionarily conserved subcomplexes, the Nup93 complex, is a major structural component linking the NPC to the membranes of the NE. Using in vitro nuclear assembly assays, we show that two components of the Nup93 complex, Nup188 and Nup205, are dispensable for NPC formation. However, nuclei lacking Nup188 increase in size by several fold compared with wild type. We demonstrate that this phenotype is caused by an accelerated translocation of integral membrane proteins through NPCs, suggesting that Nup188 confines the passage of membrane proteins and is thus crucial for the homeostasis of the different nuclear membranes.     

Functional polymorphism -31C/G in the promoter of BIRC5 gene and risk of nasopharyngeal carcinoma among chinese

Background

Baculoviral inhibitor of apoptosis repeat-containing 5 (BIRC5, also called as survivin) is a member of the inhibitor of apoptosis protein (IAP) family, which plays an important role in the occurrence and progression of cancer. Recently, a polymorphism in the promoter of BIRC5, -31C/G (rs9904341), was shown to influence BIRC5 expression.

Methods

We examined whether the -31C/G was related to the risk of developing nasopharyngeal carcinoma (NPC) in a case-control population from Guangxi province in southern China, which consists of 855 patients with NPC and 1036 controls. This polymorphism was genotyped by TaqMan assay. The genetic associations with the occurrence and progression of NPC were estimated by logistic regression.

Results

We observed a statistically significant increased occurrence of NPC associated with the CC genotype (odds ratio [OR], 1.40; 95% confidence interval [CI], 1.13–1.73; P = 0.0020) compared with the genotypes containing G allele (CG + GG genotype). However, no significant association was observed for the -31C/G with the severity of NPC (as measured by tumor-node-metastasis staging system).

Conclusion

Our findings suggest that the functional polymorphism -31C/G in the promoter of BIRC5 gene may play a role in mediating the susceptibility to NPC among Chinese.     

Lowered risk of nasopharyngeal carcinoma and intake of plant vitamin, fresh fish, green tea and coffee: a case-control study in taiwan

Background

A case-control study was conducted to evaluate the role of adult diet on nasopharyngeal carcinoma (NPC) in Taiwan.

Methods

A total of 375 incident NPC cases and 327 controls matched to the cases on sex, age, and residence were recruited between July 1991 and December 1994. A structured questionnaire inquiring complete dietary history, socio-demographic characteristics, and other potential confounding factors was used in the personal interview. Unconditional logistic regression analysis was used to estimate multivariate-adjusted odds ratio (OR_adj) with 95% confidence interval (CI) after accounting for known risk factors.

Results

Fresh fish (OR_adj, 0.56; 95% CI, 0.38–0.83 for the highest vs. lowest tertile of intake), green tea (OR_adj, 0.61; 95% CI, 0.40–0.91 for drinking ≥1 times/week vs. never) and coffee (OR_adj, 0.56; 95% CI, 0.37–0.85 for drinking ≥0.5 times/week vs. never) were inversely associated with the NPC risk. No association with NPC risk was observed for the intake of meats, salted fish, fresh vegetables, fruits and milk. Intake of vitamin A from plant sources was associated with a decreased NPC risk (OR_adj, 0.62; 95% CI, 0.41–0.94 for the highest vs. lowest tertile).

Conclusion

The study findings suggest that certain adult dietary patterns might protect against the development of NPC.     

Cholesterol Pathways Affected by Small Molecules That Decrease Sterol Levels in Niemann-Pick Type C Mutant Cells

Background

Niemann-Pick type C (NPC) disease is a genetically inherited multi-lipid storage disorder with impaired efflux of cholesterol from lysosomal storage organelles.

Methodology/Principal Findings

The effect of screen-selected cholesterol lowering compounds on the major sterol pathways was studied in CT60 mutant CHO cells lacking NPC1 protein. Each of the selected chemicals decreases cholesterol in the lysosomal storage organelles of NPC1 mutant cells through one or more of the following mechanisms: increased cholesterol efflux from the cell, decreased uptake of low-density lipoproteins, and/or increased levels of cholesteryl esters. Several chemicals promote efflux of cholesterol to extracellular acceptors in both non-NPC and NPC1 mutant cells. The uptake of low-density lipoprotein-derived cholesterol is inhibited by some of the studied compounds.

Conclusions/Significance

Results herein provide the information for prioritized further studies in identifying molecular targets of the chemicals. This approach proved successful in the identification of seven chemicals as novel inhibitors of lysosomal acid lipase (Rosenbaum et al, Biochim. Biophys. Acta. 2009, 1791:1155–1165).     

Circulating miR-17, miR-20a,miR-29c,and miR-223 Combined as Non-Invasive Biomarkers in Nasopharyngeal Carcinoma

Background

MicroRNAs have been considered as a kind of potential novel biomarker for cancer detection due to their remarkable stability in the blood and the characteristics of their expression profile in many diseases.

Methods

We performed microarray-based serum miRNA profiling on the serum of twenty nasopharyngeal carcinoma patients at diagnosis along with 20 non-cancerous individuals as controls. This was followed by a real-time quantitative Polymerase Chain Reaction (RT-qPCR) in a separate cohort of thirty patients with nasopharyngeal carcinoma and thirty age- matched non-cancerous volunteers. A model for diagnosis was established by a conversion of mathematical calculation formula which has been validated by analyzing 74 cases of patients with nasopharyngeal carcinoma and 57 cases of non-cancerous volunteers.

Results

The profiles showed that 39 and 17 miRNAs are exclusively expressed in the serum of non-cancerous volunteers and of patients with nasopharyngeal carcinoma respectively. 4 miRNAs including miR-17, miR-20a, miR-29c, and miR-223 were found to be expressed differentially in the serum of NPC compared with that of non-cancerous control. Based on this, a diagnosis equation with Ct difference method has been established to distinguish NPC cases and non-cancerous controls and validated with high sensitivity and specificity.

Conclusions

We demonstrate that the serum miRNA-based biomarker model become a novel tool for NPC detection. The circulating 4-miRNA-based method may provide a novel strategy for NPC diagnosis.     

Chemotherapeutic effect of a novel temozolomide analog on nasopharyngeal carcinoma in vitro and in vivo

Background

Many patients with nasopharyngeal carcinoma (NPC) face poor prognosis. Due to its hidden anatomical location, the tumor is usually diagnosed quite late, and despite initially successful treatment with radiation and cisplatin, many patients will relapse and succumb to the disease. New treatment options are urgently needed. We have performed preclinical studies to evaluate the potential NPC therapeutic activity of a newly developed analog of temozolomide (TMZ), an alkylating agent that is the current chemotherapeutic standard of care for patients with malignant glioma.

Results

TMZ was covalently conjugated to the natural monoterpene perillyl alcohol (POH), creating the novel fusion compound NEO212. Its impact on two NPC cell lines was studied through colony formation assays, cell death ELISA, immunoblots, and in vivo testing in tumor-bearing mice. In vitro, NEO212 effectively triggered tumor cell death, and its potency was significantly greater than that of its individual components, TMZ or POH alone. Intriguingly, merely mixing TMZ with POH also was unable to achieve the superior potency of the conjugated compound NEO212. Treatment of NPC cells with NEO212 inactivated the chemoprotective DNA repair protein MGMT (O6-methylguanine methyltransferase), resulting in significant chemosensitization of cells to a second round of drug treatment. When tested in vivo, NEO212 reduced tumor growth in treated animals.

Conclusion

Our results demonstrate anticancer activity of NEO212 in preclinical NPC models, suggesting that this novel compound should be evaluated further for the treatment of patients with NPC.

Electronic supplementary material

The online version of this article (doi:10.1186/s12929-015-0175-6) contains supplementary material, which is available to authorized users.     

Factors Predict Prolonged Wait Time and Longer Duration of Radiotherapy in Patients with Nasopharyngeal Carcinoma: A Multilevel Analysis

Purpose

Radiotherapy with or without chemotherapy is the primary treatment for patients with nasopharyngeal carcinoma (NPC). It wastes time from diagnosis to treatment. Treatment time of radiotherapy generally takes at least seven weeks. The current study aimed to evaluate factors associated with prolonged wait time and longer duration of radiotherapy in NPC patients.

Methods and Materials

From Taiwan''s National Health Insurance research database, we identified 3,605 NPC patients treated with radiotherapy between 2008 and 2011. Wait time was calculated from the date of diagnosis to the start of radiotherapy. The impact of each variable on wait time and duration of radiotherapy was examined by multilevel analysis using a random-intercept model.

Results

The mean wait time and duration of radiotherapy were 1.78±3.33 and 9.72±7.27 weeks, respectively. Multilevel analysis revealed prolonged wait time in patients aged 45–65 years, those receiving radiotherapy alone, those with more comorbidities, those with low SES, and those living in eastern Taiwan. A prolonged duration of radiotherapy was associated with receipt of concurrent chemoradiotherapy, more comorbidities, and moderate SES.

Conclusions

Understanding the factors associated with longer wait times and duration of radiotherapy in patients with NPC may help healthcare providers better assist both these patients and potentially those with other head-and-neck cancers.     

Diabetes,Prediabetes and the Survival of Nasopharyngeal Carcinoma: A Study of 5,860 Patients

Background

The incidence of diabetes is increasing. But the impact of diabetes and prediabetes on survival of patients with nasopharyngeal carcinoma (NPC) has received little evaluation.

Methods

In a cohort of 5,860 patients, we compared the disease specific survival (DSS), locoregional relapse-free survival (LRFS) and distant metastasis-free survival (DMFS) of patients with diabetes, prediabetes and normoglycemia defined by pretreatment fasting plasma glucose (FPG) using Kaplan–Meier method, log-rank test and Cox proportional hazards model.

Results

Comparing to normoglycemic patients, the diabetic and the prediabetic were generally older, fatter, had hypertension, heart diseases and hyperlipaemia and usually received radiotherapy alone. But both the diabetic and the prediabetic had similar DSS, LRFS and DMFS to normoglycemic patients, even adjusting for such important factors as age, gender, smoking, drinking, hypertension, heart diseases, body mass index, hyperlipaemia, titer of VCA-IgA and EA-IgA, pathology, T-stage, N-stage, chemotherapy and radiotherapy (P>0.05 for all). Additionally, the findings remained unchanged in sensitivity analysis by excluding patients with known diabetes history and in subgroups of the various factors.

Conclusions

The diabetic and prediabetic NPC patients had similar survival to normoglycemic NPC patients. These data, in the largest reported cohort, are the first to evaluate the association between diabetes, prediabetes and the survival in NPC. The findings are relevant to patient management and provided evidence of the effect on this disease exerted by comorbidities.     

Evaluation of an anti-tumor necrosis factor therapeutic in a mouse model of Niemann-Pick C liver disease

Background

Niemann-Pick type C (NPC) disease is a lysosomal storage disease characterized by the accumulation of cholesterol and glycosphingolipids. The majority of NPC patients die in their teen years due to progressive neurodegeneration; however, half of NPC patients also suffer from cholestasis, prolonged jaundice, and hepatosplenomegaly. We previously showed that a key mediator of NPC liver disease is tumor necrosis factor (TNF) α, which is involved in both proinflammatory and apoptotic signaling cascades. In this study, we tested the hypothesis that blocking TNF action with an anti-TNF monoclonal antibody (CNTO5048) will slow the progression of NPC liver disease.

Methodology/Principal Findings

Treatment of wild-type C57BL/6 mice with NPC1-specific antisense oligonucleotides led to knockdown of NPC1 protein expression in the liver. This caused classical symptoms of NPC liver disease, including hepatic cholesterol accumulation, hepatomegaly, elevated serum liver enzymes, and lipid laden macrophage accumulation. In addition, there was a significant increase in the number of apoptotic cells and a proliferation of stellate cells. Concurrent treatment of NPC1 knockdown mice with anti-TNF had no effect on the primary lipid storage or accumulation of lipid-laden macrophages. However, anti-TNF treatment slightly blunted the increase in hepatic apoptosis and stellate cell activation that was seen with NPC1 knockdown.

Conclusions/Significance

Current therapeutic options for NPC disease are limited. Our results provide proof of principle that pharmacologically blocking the TNF-α inflammatory cascade can slightly reduce certain markers of NPC disease. Small molecule inhibitors of TNF that penetrate tissues and cross the blood-brain barrier may prove even more beneficial.